Overexpression of USF increases TGF-beta1 protein levels, but G1 phase arrest was not induced in FRTL-5 cells.
Transforming growth factor-beta1 (TGF-beta1) is a potent inhibitor of cellular growth and proliferation by G1 phase arrest or apoptosis. We investigated the association of TGF-beta1 with the anti-proliferative effect of upstream stimulatory factor (USF) in Fischer rat thyroid cell line (FRTL-5) cells. [methyl-(3)H] thymidine uptake was measured after treatment of FRTL-5 cells with TGF-beta1 to identify its anti-proliferative effect. USF-1 and USF-2 proteins were in vitro translated, and an electrophoretic mobility shift assay was performed to identify the interaction between USF and the TGF-beta1 promoter. FRTL-5 cells were transfected with USF cDNA, and then the expression of TGF-beta1 was examined with Northern and Western blotting. The cell cycle-regulating proteins associated with TGF-beta1 were also measured. TGF-beta1 significantly inhibited [methyl-(3)H] thymidine uptake in FRTL-5 cells. Two specific binding sites for USF were found in the TGF-beta1 promoter: -1,846 approximately -1,841 (CACATG) and -621 approximately -616 (CATGTG). Overexpression of USF increased both the mRNA levels and protein levels of TGF-beta1. However, the expression of cyclin D1, CDK4, cyclin E, and CDK2, and the phosphorylation of retinoblastoma protein remained unchanged. Overexpression of USF in FRTL-5 cells increased the expression of TGF-beta10 through specific binding to TGF-beta1 promoter. However, the USF-induced expression of TGF-beta1 did not cause G1 arrest.
Kim
Keun-Sook
KS
Department of Medicine, Samsung Medical Center, Jungkyunkwan University School of Medicine, Seoul, Korea.
Jung
Hye Seung
HS
Chung
Yun Jae
YJ
Jung
Tae Sik
TS
Jang
Hye Won
HW
Lee
Myung-Shik
MS
Kim
Kwang-Won
KW
Chung
Jae Hoon
JH
D000818:Animals; D017209:Apoptosis; D001665:Binding Sites; D002453:Cell Cycle; D002460:Cell Line; D016193:G1 Phase; D005786:Gene Expression Regulation; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D051381:Rats; D013936:Thymidine / Q000737:chemistry; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D051720:Upstream Stimulatory Factors / Q000378:metabolism*
D053773:Transforming Growth Factor beta1; D051720:Upstream Stimulatory Factors; C497182:Usf1 protein, rat; C497187:Usf2 protein, rat; D013936:Thymidine